We have been engaged in studies of SV40 vectors for molecular cloning in mammalian cells. These methods are being developed for molecular cloning of well defined eukaryotic gene regions which have previously been cloned in plasmid or phage lambda vectors of E. coli. In addition, work has been undertaken on different types of SV40 vectors used for the transduction of segments of phage lambda DNA as well as specific regions of E. coli DNA into simian or rodent cells. At present, this work is progressing under P3 physical containment at UCLA. Previously we have propagated a suppressor tRNA gene of E. coli in monkey cells utilizing a vector carrying the early gene region of SV50 in addition to the origin for DNA replication. The hybrid DNA was propagated by mixed infection of monkey cells with an early mutant (tsA) helper. Very recently in studies with this SV40-su ions III hybrid DNA we have discovered that two potentially promising helper virus-free cloning approaches are feasible with defective SV40 vectors of this type. In these approaches either permissive or non-permissive cells or multiplication by SV40 are infected with purified hybrid DNA. Subsequently cloned lines of infected cells are isolated and screened for the presence of hybrid DNA sequences. In the case of the permissive cell lines, such as green monkey or rhesus monkey kidnay cells, we have found that viable cell populations can be obtained which carry substantial quantities of free replicating hybrid DNA. In the continuation of these recombinant DNA studies, we propose to explore the fate of specific histone genes from the lower eukaryotic organism, sea urchin after linkage of these genes to SV40 vectors. We will both propagate the hybrids in permissive monkey kidney cells using appropriate early and late SV40 mutants as helpers and also select permissive cells transfected with the purified hybrid DNA. We will then go on to examine whether specific transcripts of the histone gene region linked to the SV40 vector can be identified and then attempts will be made to determine whether production of the histone proteins are, in fact, taking place.